【摘要】 目的 研究N-乙酰半胱氨酸酰胺（NACA）对脂多糖（LPS）诱导的RAW264.7巨噬细胞炎症的保护作用及作用机制，为NACA治疗脓毒症提供理论依据。方法 CCΚ-8法检测NACA和N-乙酰半胱氨酸（NAC）对RAW264.7细胞活力的影响；采用ELISA检测NACA和NAC对LPS诱导的RAW264.7细胞上清中TNF-α和IL-6水平；qRT-PCR检测NACA对LPS诱导的RAW264.7细胞中TNF-α、IL-6、IL-1β、ATF4、CHOP的mRNA表达的影响；Western blot检测NACA对LPS诱导的RAW264.7细胞中TLR4/NF-κB及PERΚ/ATF4/CHOP信号通路中关键蛋白表达的影响。结果 0.1～5 mmol/L的NACA和NAC对RAW264.7细胞活力无明显的影响（P>0.05）。1 mmol/L和5 mmol/L的NACA和NAC可显著降低LPS诱导的RAW264.7细胞上清中TNF-α和IL-6的水平（P<0.01），且NACA的降低效果优于统计量的NAC。1 mmol/L和5 mmol/L的NACA也可显著降低LPS诱导的RAW264.7细胞中TNF-α、IL-6、IL-1β、ATF4、CHOP mRNA的表达（P<0.05、P<0.01）。Western blot结果表明，1 mmol/L和5 mmol/L的NACA可显著降低TLR4、p-IκB、p-65、p-PERK、p-eIF2α、ATF4、Bip和CHOP蛋白的表达（P<0.05、P<0.01）。结论 NACA可能通过TLR4/NF-κB及PERΚ/ATF4/CHOP信号通路抑制LPS诱导的RAW264.7巨噬细胞的炎症反应，为NACA治疗脓毒症的抗炎性药物筛选提供了实验依据。更多还原

【Abstract】 This study was performed to investigate the protective effects of N-acetylcysteine amide(NACA) on RAW264.7 macrophages from lipopolysaccharide(LPS)-induced inflammation and the mechanism of action, and to provide a theoretical basis for the treatment of sepsis with NACA. CCΚ-8 assay was used to detect the effects of NACA and N-acetylcysteine(NAC) on the viability of RAW264.7 cells; ELISA was used to detect the effects of NACA and NAC on LPS-induced TNF-α and IL-6levels in the supernatant of RAW264.7 cells; qRT-PCR was used to detect the effects of NACA on LPS-induced TNF-α, IL-6, IL-1β, ATF4, CHOP mRNA expression in RAW264.7 cells. q RT-PCR was performed to detect the effect of NACA on the expression of key proteins in TLR4/NF-κB and PERΚ/ATF4/CHOP signaling pathways in LPS-induced RAW264.7 cells. Data showed that NACA and NAC at 0.1 to 5 mmol/L had no significant effect on RAW264.7 cell viability(P>0.05). 1 mmol/L and 5 mmo/L of NACA and NAC significantly reduced the levels of TNF-α and IL-6 in the supernatant of LPS-induced RAW264.7 cells(P<0.01), and the reduction induced by NACA was better than the statistical amount of NAC. 1 mmol/L and 5 mmol/L NACA also significantly reduced the expression of TNF-α, IL-6, IL-1β, ATF4, and CHOP mRNA in LPS-induced RAW264.7 cells(P<0.05, P<0.01).Western blot results showed that 1 mmol/L and 5 mmol/L NACA significantly reduced the expression of TLR4, pIκB, p65, p-PERK, p-eIF2α, ATF4, Bip and CHOP proteins(P<0.05, P<0.01). Taken together, NACA may inhibit LPS-induced inflammatory responses in RAW264.7 macrophages via TLR4/NF-κB and PERΚ/ATF4/CHOP signaling pathways, providing an experimental basis for anti-inflammatory drug screening of NACA in the treatment of sepsis.更多还原